Innate immune cells, which act as a first line of defense against disease-causing pathogens, mount responses upon detection of microbial structural patterns conserved on a wide array of bacteria, viruses and fungi. Effective innate responses rid an individual of a vast number of invading pathogens and also appear vital for optimal stimulation of B and T lymphocyte memory responses. On the other hand, overactive innate responses are correlated with allergic and autoimmune diseases. Thus, a basic understanding of innate immune responses and the microbial pattern recognition machinery underlying these responses is crucial to development of new therapies for infectious and autoimmune diseases. In addition, insights into innate immune stimulation of memory lymphocytes can assist in development of effective vaccines. The microbial pattern recognition machinery used by innate immune systems to detect pathogens appears highly conserved, and for this reason, the study of immunity in simple animals such as Drosophila has led to the discovery and characterization of many microbial pattern recognition mechanisms, notably the Toll-like receptors. The study of the molecular basis of the innate immune system's recognition of microbes would be facilitated by the use of a simple eukaryotic cell system with powerful tools for genetic analysis. Here we propose to explore the possibility that the social amoeba uses the same types of microbial pattern recognition machinery to detect bacterial prey and defend against bacterial pathogens as do mammalian innate immune cells and would thus provide such a model. Indeed, D. discoideum has proven to be a useful model for the study of chemotactic responses by mammalian immune cells, and the parallels in the signaling pathways that mediate chemotaxis in these evolutionary diverse cells are remarkable. Several lines of evidence suggest that D. discoideum uses conserved pattern recognition mechanisms. Its genome encodes for proteins homologous to known pattern recognition molecules, and analysis of some of these proteins has indicated a role in bacterial recognition. In addition, our preliminary results show that D. discoideum initiates cellular responses upon detection of particular microbial patterns. To test our hypothesis that microbial pattern recognition machinery is conserved in D. discoideum we plan to 1) further analyze D. discoideum responses to bacterial patterns, 2) use knockout and overexpression technologies to characterize D. discoideum proteins homologous to known pattern recognition molecules, and 3) employ mutant screening and yeast-two-hybrid techniques to identify new D. discoideum proteins involved in bacterial recognition. The results from our studies should allow further characterization of microbial pattern recognition in D. discoideum. Given the conservation of microbial pattern recognition mechanisms among a wide array of species, our findings should also give insight into pattern recognition in mammalian innate immune systems. PUBLIC HEALTH RELEVANCE: Innate immune responses are critical both for the quick and efficient removal of most pathogens and for the stimulation of effective long-term memory responses from T and B lymphocytes. Conversely, overactive innate responses are associated with allergic and autoimmune diseases. A greater understanding of the molecular mechanisms underlying microbial recognition by innate immune systems could allow for the regulation of innate immune responses in infectious and autoimmune diseases and allergies. Insights into the stimulation of long-term memory responses by innate immune systems should also allow for design of more effective vaccine strategies. Our proposal here to use the social amoeba Dictyostelium discoideum as a model system to study the molecular mechanisms underlying microbial pattern recognition in innate immunity should lend insight into mechanisms that may be conserved in mammalian immune systems and may inform development of new therapeutics for infectious and autoimmune diseases.